WEEK 7 LABORATORYPHYLOGENETIC ANALYSIS OF ASTERID EUDICOTS |
Families sampled Sampling characters Character list Cladistic analysis Phenetic analysis Report
I. AIMS
This laboratory exercise involves a phylogenetic analysis of the Asterid eudicots. The asterids are a fairly natural group of eudicots best identified by the fusion of petals to form a corolla tube. The aims of this exercise are (1) to familiarize yourself with the procedures of character selection and data collection; i.e., find and score as many variable characters as possible among the representative asterid families; (2) to allow you to use computer programs that can analyze such data with both phenetic and cladistic methods to determine relationships among the asterid families; and (3) to give you the opportunity to make taxonomic decisions based on your results. See lecture notes and handouts for review.
A. Operational Taxonomic Units (OTUs or families investigated)
Fourteen genera representing 14 different families of Asterids, and one genus representing the Rosids (to be used as the outgroups) are provided at each table or as demonstrations at the front desk (see Plant Systematics [T], Zomlefer[Z] and Cronquist [C] for information on each family/genus). Many of the genera are tropical or subtropical so you will not find them in Gleason & Cronquist. For these (and others), consult Cronquist's An Integrated System of Classification of Flowering Plants [C]; and Heywood's Flowering Plants of the World at the front. Although not providing information about these genera, these references will provide considerable detail concerning the families. For additional information on these families, go to the family links in the University of Wisconsin Plant Systematics Collection homepage. The recent ordinal molecular phylogeny of Asterids is largely reflected in the APG system of classification shown below (see also the asterid family molecular phylogeny).
Asterid Eudicots Order Gentianales Gentianaceae 1. Exacum 305 [T] 244 [Z] 871 [C] Apocynaceae 2. Allamanda 301 [T] 239 [Z] 876 [C] Rubiaceae 3. Pentas 305 [T] 236 [Z] 995 [C] Order Solanales Solanaceae 4. Petunia 319 [T] 213 [Z] 892 [C] Convolvulaceae 5. Ipomoea 319 [T] 215 [Z] 895 [C] Order Lamiales Lamiaceae 6. Salvia 308 [T] 265 [Z] 924 [C] Verbenaceae 7. Lantana --- 262 [Z] 920 [C] Scrophulariaceae 8. Antirrhinum 315 [T] * 250 [Z] 951 [C] Bignoniaceae 9. Tecomaria 308 [T] 248 [Z] 968 [C] Acanthaceae 10. Justicia 308 [T] 259 [Z] 963 [C] Order Asterales Campanulaceae 11. Campanula --- 211 [Z] 983 [C] Lobeliaceae 12. Lobelia --- # 211 [Z] 983 [C] Goodeniaceae 13. Scaevola 331 [T] --- 989 [C] Order Ericales Ericaceae 14. Erica 293 [T] 77 [Z] 479 [C] Order Myrtales --- Rosid [for outgroup purposes] Onagraceae 15. Fuchsia 259 [T] 229 [Z] 645 [C] * now placed in family Plantaginaceae
# now placed in family Campanulaceae
B. Characters and Character States
Your group will examine the living plants of each family and look for vegetative, floral, and fruit characters that vary among the families. You have been provided already a list of characters; the character states that the fifteen families possess can be obtained by (1) working in groups; (2) examining the plants macroscopically; (3) examining the flowers/fruits microscopically; (4) looking at descriptions of the families and/or genera in your Text, Zomlefer, Cronquist, and other texts provided at the front of the lab; and (5) being persistent and inquisitive. Additional characters, if necessary, can be obtained from various literature sources (Text, Zomlefer, Cronquist, Heywood).
Only qualitative characters (presence/absence; number of sepals; color; shape; etc.) should be scored on the provided blank data matrix sheets. Please do not use quantitative characters that vary continuously (length/width of leaves; number of flowers; etc.). We will avoid these latter quantitative characters as they are difficult to use in a cladistic fashion, although they can be handled in phenetic analyses.
Make certain you are carefully distinguishing between characters and character states. Character states that can not be determined for certain families (lacking information totally e.g., do not have fruits or can not find the relevant information on fruits for the family; or inapplicable to that taxa e.g., shape of corolla tube in family without fusion of petals) should be treated as "?" in your matrix. Otherwise, give each character state an integer value (0 and 1 for binary character states; 0, 1, 2 for characters with 3 states; etc.).
Once your group has completed the data matrix, enter the data into a file using the program MacClade on the computer in the lab. Open the "Phylogenetics Lab" folder in the center of the desktop, and drag the file "Asterids.template" on the desktop into the icon "MacClade". [If a message about fonts is encountered, just press "return".] When the file opens up, you can enter "0", "1", "2" etc. into the appropriate cell (columns are the characters, rows are the taxa). Use the arrow keys at bottom left to speed up the data entry procedure. You can make new columns for additional characters you want to include or delete present characters by going to the "Edit" menu at top (see TA for further instructions). After you have entered some of the data, go to "FILE" menu at top left and "SAVE FILE AS" your file as "Asterids.[your last name]" making sure that it is saved within the folder "Phylogenetics Lab". Save again when you are completely done entering the data.
C. Cladistic Analysis [see lecture notes for discussion of cladistics]
The cladistic computer program "Phylogenetic Analysis Using Parsimony" (PAUP* 4.0) on the laboratory computer will be used to produce a genealogy or cladogram of the Asterids based on your morphological data set. Someone from your group should act as leader to do this part; it would be best, however, if your whole group could watch as the computer program is being used. You might need to set a time over the next week to come in and use the program.
Open the "Phylogenetics Lab" folder and drag your file "Asterids.[your last name]" into the icon "Paup 4.0" on the desktop. Then press the keys "apple" + "R" at the same time to run the program with your data set. The outgroup is already indicated in the file. The basic cladistic run is performed by going to the "Analysis" menu at top right (second from right), and indicating "Heuristic Search". All the default conditions are in place, so simply click the "Search" button. Alternatively, one can perform "Branch and Bound Search" under the "Analysis" menu. When the search is done, one or more trees will be found: click the "Close" button to get out of the searching mode. Save the tree or trees by going to the "Trees" menu at extreme top right and indicating "Save Trees to File" and name your treefile "Asterids.[your name].tree" and saving it to the "Phylogenetics Lab" folder.
If a single tree is found, go to the "Trees" menu, indicate "Print Trees", click on the "Print" button a printout of the tree will be obtained in Room 258, and then exit the "Print Trees" mode by clicking "Done" (you can preview the tree before printing by clicking the "Preview" button just above the "Print" button) . If more than one tree was found, you can make a consensus tree by going to the "Trees" menu and indicating "Compute Consensus". Follow the default conditions and click "OK". To print out the consensus tree, go to the "Trees" menu and indicate "Print Consensus Tree".
D. Phenetic Analysis [see lecture notes for discussion of phenetics]
Phenetic analysis will be done with computer program PAUP * 4.0 as well. You will try two different phenetic analyses: Neighbor-joining (NJ) and UPGMA. The latter, but not the former, uses an assumption of equal rates of evolution along each branch. Therefore, UPGMA will attempt to derive a tree that is more nearly symmetrical in terms of character changes from any point on the tree to any two terminal tips (in our case, genera). NJ will attempt to find the tree that is most consistent with the distance values for each pair wise comparsions without this added assumption. Unlike cladistic analyses, both of these distance approaches will each generate a single tree.
Open the "Phylogenetics Lab" folder and drag your file "Asterids.[your last name]" into the icon "Paup 4.0" on the desktop. Then press the keys "apple" + "R" at the same time to run the program with your data set. The outgroups are already indicated in the file. The distance based analyses are performed by going to the "Analysis" menu at top right (second from right), and indicating "Neighbor Joining/UPGMA". All the default conditions are in place, so simply indicate either "Neighbor joining" or "UPGMA" in the upper left "Algorithm" box, and then press the "OK" button. When the search is done, the single best tree will scroll through your screen.
To print the tree, go to the "Trees" menu, indicate "Print NJ Tree" (or "Print UPGMA Tree"), click on the "Print" button a printout of the tree will be obtained in Room 258, and then exit the "Print Trees" mode by clicking "Done" (you can preview the tree before printing by clicking the "Preview" button just above the "Print" button). Save the tree by going to the "Trees" menu at extreme top right and indicating "Save Trees to File" and name your treefile "Asterids.[your name].NJ" or "Asterids.[your name].UPGMA" and saving both to the "Phylogenetics Lab" folder.
Each student will work in a small group to obtain the original character state matrix and to do the cladistic and phenetic analyses. Each student, however, should work independently on the report and will hand in his/her own report in two weeks (due during lab of Oct. 26-28). This report will be part of your laboratory (20 pts.) and should include minimally (1) the data matrix, (2) the cladogram or consensus cladogram, (3) the NJ and UPGMA trees, and (4) several paragraphs on "Taxonomic Analysis".
The "Taxonomic Analysis" section should compare the different trees (cladistic and phenetic) with the new APG Classification (handout after Classification lectures). Questions to ponder and discuss include: (1) Are the families placed in the orders according to the APG classification system and to the recent molecular phylogeny of asterids? (2) Based on your data, what character(s) - when mapped onto the APG or molecular phylogeny - support different branches? (3) What reclassification would you suggest based on your analyses? or are there problems with your data or analysis that would prevent you from making any changes now?
Instructions for mapping your data set onto the APG/Soltis Tree
1. Open your datafile (Asterids.[your last name]) in MacClade
2. Open the Tree Window (apple + T).
3. In Tree Window, go to "Tree" menu, and "Open Tree File." Open "AsteridsLab.2000.tre." A shortened version of the APG tree should appear
4. Go to the "Trace" menu, choose "Trace Character." The tree should now be colored, a little box at the lower right hand corner of the screen should show which character is currently being traced, with different colors showing the character states at different branches. Note: zebra striping means that the character is "equivocal" on that particular branch segment. This means that several character states are equally probable at this point.
5. To answer the question posed in the lab writeup, you need to determine which of your characters show less homoplasy (on the APG tree) - are derived fewer times. Spend more time discussing the more useful.
